The ultimate goal of this project is the development of a novel therapeutic strategy for the treatment of cancer capitalizing on the unique antiproliferative effects of the alkaloid lycorine on apoptosis resistant cancers. Although a large majority of current cancer drugs exhibit their effects through the induction of apoptosis in tumor cells, many types of cancer, such as glioma, melanoma, NSCLC, among others, are resistant to proapoptotic stimuli and are typically associated with dismal prognoses. Therefore, new chemotypes that display antiproliferative effects against such apoptosis-resistant cancers and/or inhibit tumor metastatic spread are urgently needed. In preliminary work, the PI's collaborative chemical biology/drug discovery team has shown that the alkaloid lycorine modifies actin cytoskeleton organization in cancer cells, and impairs both their proliferation and migration. Lycorine exerts these effects with both apoptotic-resistant and susceptible cell lines, sensitizes cancer cells to proapoptotic agents, and displays 15 times greater potency toward cancer versus normal cells. Furthermore, lycorine showed significant therapeutic benefit in vivo in the B16F10 mouse melanoma model, consisting of orthotopically brain-grafted melanoma cells. These preliminary results form a compelling case for a thorough understanding of lycorine's mode of action in cancer cells and utilization of this knowledge for the development of novel strategies to treat cancers with dismal prognoses. The proposed work involves the preparation of lycorine- alkyne probes, which will be evaluated against a panel of apoptosis-resistant cancer cells with the goal of identifying derivatives retaining the antiproliferative potency and mode of action of lycorine. Suitable alkyne probes will then be used in protein pull-down assays, aimed at target protein identification. Several back-up target identification and target validation strategies are also proposed to confirm this innovative chemotherapeutic approach.